In vivo behaviour of a biodegradable poly(trimethylenecarbonate) barrier membrane: a histological study in rats

A. C. Van Leeuwen • T. G. Van Kooten •

D. W. Grijpma • R. R. M. Bos

Received: 13 December 2011 / Accepted: 24 April 2012 / Published online: 9 May 2012

� The Author(s) 2012. This article is published with open access at Springerlink.com

Abstract The aim of the present study was to evaluate

the response of surrounding tissues to newly developed

poly(trimethylene carbonate) (PTMC) membranes. Fur-

thermore, the tissue formation beneath and the space

maintaining properties of the PTMC membrane were

evaluated. Results were compared with a collagen mem-

brane (Geistlich BioGide), which served as control. Single-

sided standardized 5.0 mm circular bicortical defects were

created in the mandibular angle of rats. Defects were

covered with either the PTMC membrane or a collagen

membrane. After 2, 4 and 12 weeks rats were sacrificed

and histology was performed. The PTMC membranes

induced a mild tissue reaction corresponding to a normal

foreign body reaction. The PTMC membranes showed

minimal cellular capsule formation and showed signs of a

surface erosion process. Bone tissue formed beneath the

PTMC membranes comparable to that beneath the collagen

membranes. The space maintaining properties of the

PTMC membranes were superior to those of the collagen

membrane. Newly developed PTMC membranes can be

used with success as barrier membranes in critical size rat

mandibular defects.

1 Introduction

Guided bone regeneration (GBR) is a widely used modality

for restoring bone deficiencies. In GBR the use of barrier

membranes leads to predictable bone formation, by pre-

venting in-growth of fibroblasts and provision of space for

osteogenesis within a blood clot formed in the defect [1]. A

variety of biocompatible membranes have been used to

achieve this desired barrier effect. The optimal barrier

membrane should exert biocompatible, synthetic, degrad-

able and space maintaining properties [2, 3]. Currently,

non-resorbable membranes have better space maintaining

properties compared to resorbable membranes. However, a

major disadvantage of non-resorbable membranes is the

need for their removal in a second operation.

The majority of clinically used resorbable membranes

are based on collagen. As collagen is an animal derived

product, these membranes carry the risk of disease trans-

mission from animal to human [3–5]. Other available

resorbable barrier membranes are synthetic polymeric

membranes based on lactide and glycolide. However, due to

an extensive foreign body reaction, adverse effects like

postoperative swelling have been reported when using these

materials [6–13]. It is known that these materials can produce

significant amounts of acidic compounds during degradation

in the body, and since bone dissolves in acidic environments,

it can be expected that these polymers will not be the most

suited materials for use in GBR [7, 12, 14, 15].

Recently, we have developed a flexible synthetic

biodegradable barrier membrane prepared from poly(tri-

methylene carbonate) (PTMC) for GBR [16]. PTMC is a

A. C. Van Leeuwen (&) � R. R. M. Bos

Department of Oral and Maxillofacial Surgery,

University Medical Center Groningen, PO Box 30001,

9700 RB Groningen, The Netherlands

e-mail: a.van.leeuwen01@umcg.nl

T. G. Van Kooten � D. W. Grijpma

Department of Biomedical Engineering, University Medical

Center Groningen and University of Groningen, PO Box 196,

9700 AD Groningen, The Netherlands

D. W. Grijpma

MIRA Institute for Biomedical Engineering and Technical

Medicine, and Department of Biomaterials Science and

Technology, Faculty of Science and Technology, University of

Twente, PO Box 217, 7500 AE Enschede, The Netherlands

123

J Mater Sci: Mater Med (2012) 23:1951–1959

DOI 10.1007/s10856-012-4663-x

flexible, rubber-like, amorphous and biodegradable poly-

mer. The monomer from which it is prepared, trimethylene

carbonate (TMC), has been used to prepare copolymers

for use in barrier membranes in the medical field before

[17–20]. By gamma irradiation under vacuum, form-stable

elastomeric networks can be formed [21]. In vitro and in

vivo research has shown that this polymer is both bio-

compatible and can be degraded by surface erosion without

the formation of acidic degradation products [22, 23].

In the aforementioned study [16] the PTMC membrane,

prepared from TMC only, was evaluated and compared

with a collagen (Geistlich BioGide) and an expanded-

polytetrafluoroethylene (e-PTFE, GoreTex) membrane and

a non-treated control site for its suitability as a barrier

membrane for use in GBR over bony defects in rats. Both

quantitative micro-radiographical and quantitative micro-

computed tomographical analysis showed excellent

amounts of bone formed underneath the PTMC membrane

[16]. Evaluation after 12 weeks showed that comparable

amounts of bone had formed underneath the PTMC and

reference membranes. The mean percentages of regener-

ated bone were 74, 71, 83 and 33 %, for respectively

the collagen, PTMC e-PTFE and the non-treated control

group [16].

The purpose of this current study was to evaluate his-

tologically the response of the surrounding tissue to the

PTMC membrane, the tissue formation beneath the PTMC

membrane, and the space maintaining properties of the

PTMC membrane. Furthermore, the degradation of the

membrane was assessed. The collagen membrane served as

a reference.

2 Materials and methods

2.1 Membrane synthesis

Polymerization grade 1,3-trimethylene carbonate (TMC)

was obtained from Boehringer Ingelheim, Germany.

Stannous octoate (SnOct2 from Sigma, USA) was used as

received. The used solvents were of analytical grade and

purchased from Biosolve, The Netherlands. PTMC was

synthesized by ring opening polymerization of TMC

monomer under vacuum at 130 �C for a period of three

days using stannous octoate as a catalyst. The polymer was

purified by dissolution in chloroform and precipitation into

a fivefold excess of ethanol 100 %. The precipitated PTMC

was dried under vacuum at room temperature. Analysis of

the synthesized polymer by proton nuclear magnetic reso-

nance (1H NMR), gel permeation chromatography (GPC)

and differential scanning calorimetry (DSC) according to

standardized procedures [21] indicated that high molecular

weight polymer had been synthesized. GPC measurements

showed that Mw = 443000 and Mn = 332000 g/mol, while

NMR indicated that the monomer conversion was more

than 98 %. The glass transition temperature of this amor-

phous polymer was approximately -17 �C, as thermal

analysis showed.

The purified polymer was then compression moulded at

140 �C and a pressure of 3.0 MPa (31 kg/cm2) using a

Carver model 3851-0 laboratory press (Carver Inc, USA)

into films with a thickness of 0.3 mm and a diameter of

8 mm using stainless steel moulds. The PTMC membranes

were then sealed under vacuum and exposed to 25 kGy

gamma irradiation from a 60Co source (Isotron BV, The

Netherlands). This sterilization method leads to simulta-

neous crosslinking of the polymer [21].

2.2 Study design

The gamma irradiated PTMC membranes were evaluated by

a subperiosteal implantation in rats. All procedures per-

formed on the animals were done according to international

standards on animal welfare and complied with the Animal

Research Committee of the University Medical Centre

Groningen. Twenty-four Sprague–Dawley rats from an

earlier study [16] were included in here. The surgical pro-

cedure consisted of drilling one bicortical critical size

(5.0 mm diameter) mandibular defect in the left mandibular

angle with a trephine. The created defects were covered with

a barrier membrane on the buccal and lingual side using

either the PTMC membrane, or a collagen (Geistlich Bio-

Gide, Geistlich, Switzerland) membrane which served as

reference. (The procedures are described in the section

‘‘Implantation procedure’’.) At three different time periods

(2, 4 and 12 weeks) the rats were sacrificed. Per time period

eight rats were evaluated; four rats treated with the collagen

membrane and four rats treated with the PTMC membrane.

The rats were histologically evaluated for the response of the

surrounding tissue to the membranes, and the tissue prolif-

eration at the defect site, covered by the membranes. Fur-

thermore, space maintaining properties and degradation of

the PTMC membranes were assessed.

(In a separate study, of which the results will be reported

in another communication, the bone sample that was

obtained with the trephine was transplanted to the contra-

lateral mandibular angle to evaluate the suitability of the

membranes on modelling and incorporation of autologous

bone grafts.)

2.3 Implantation procedure

The animals were anaesthetized with nitrous–oxygen–

isoflurane. After the mandibular area was shaved and

1952 J Mater Sci: Mater Med (2012) 23:1951–1959

123

disinfected, a peri angular incision was made and a

standardised circular 5.0 mm critical size bicortical defect

was drilled with a trephine in the left mandibular angle [24,

25]. In one group the PTMC membrane was used to cover

the defects. In the other group the defects were covered

with a collagen membrane (Geistlich BioGide, Geistlich,

Switzerland). The wound was closed in layers using

resorbable sutures (Vicryl Rapide 4-0, Ethicon, USA). A

single dose of Temgesic (0.05 mg/kg) was administered

perioperative for postoperative pain relief, and the diet was

composed of standard laboratory food.

At three different time intervals (2, 4 and 12 weeks), the

rats were anaesthetised by nitrous–oxygen–isoflurane

inhalation and sacrificed by an intracardially injected

overdose of pentobarbital. The mandibles were explanted

and fixed in 4 % phosphate buffered formaline solution.

2.4 Light microscopy: histological scoring analysis

After explantation, the specimens were decalcified and

dehydrated in a series of ethanol. The specimens were

embedded in glycidyl methacrylate. The tissue blocks were

cut perpendicularly to the defects with a microtome. The

resulting histological sections were 2 lm thick. From all

samples two sections were stained, one series with tolui-

dine blue and one with toluidine blue/basic fuchsin. The

histological sections were digitalized with a ScanScope GL

(Aperio Technology Inc., Vista, CA). Digital images were

then stored for further analysis. Histological sections were

qualitatively and semiquantitatively analyzed using a

modified histological scoring analysis (Table 1). Two his-

tological sections were evaluated for each animal. The

investigators were blinded for time interval and type of

material used during evaluation of the explanted samples.

During analysis each section was evaluated with respect to

the foreign body response (soft tissue response to PTMC

membrane), (the kind of) tissue proliferation at the defect

site and finally for the space maintaining properties of the

membrane. Each section received a single score. The

scores for each time interval were then averaged; mean and

standard deviation were reported.

2.5 Statistical analysis

The data sets were statistically evaluated with a Fisher’s

Exact Test using SPSS 17 (Statistical Package for the

Social Sciences, SPSS Inc., USA). The null hypothesis (the

means of each set are equal) was evaluated with 95 %

confidence level (a = 0.05).

3 Results

3.1 Clinical observations

None of the rats died or suffered from postoperative

complications. The animals started eating immediately

after the surgical procedure and did not loose weight.

3.2 Descriptive microscopic observations

A gross light microscopical inspection of all histological

sections was performed. Defects and the PTMC and col-

lagen barrier membranes could easily be identified after

2–4 weeks (Figs. 1, 2). The PTMC membrane appeared

white in the sections, the collagen barrier membrane

appeared similar to (collagen) connective tissue, although

it could be differentiated from host (collagen) connective

tissue.

At 12 weeks it was more difficult to identify the defects,

whereas in some animals the defect had closed. The col-

lagen membrane could not be identified anymore at

12 weeks because, besides extensive signs of degradation

of the membrane, the differentiation between membrane

and host collagen was virtually impossible. The PTMC

membrane showed extensive signs of degradation. Only

Table 1 Semiquantitative histological grading scale

Soft tissue response to membrane

Fibrous, mature, not dense, resembling connective

or at tissue in the noninjured regions

4

Shows blood vessels and young fibroblasts, few

macrophages and giant cells are present

3

Shows macrophages and other inflammatory cells in

abundance, but connective tissue components between

2

Dense and exclusively of inflammatory type 1

Cannot be evaluated because of infection or other

factors not necessarily related to the material

0

Tissue proliferation in defect

Mature bone and differentiation of bone marrow 4

Bone or osteoid formation 3

Fibrous connective tissue: collagen fibers at defect site 2

Fibrous connective tissue: cellular and vascular components 1

Cannot be evaluated because of infection or other

factors not necessarily related to the material

0

Space maintaining properties of membrane

No contact between membranes at defect site, bone

formation in between

4

No contact between membranes at defect site, connective tissue

in between

3

Contact between membranes at defect site, bone formation

present

2

Contact between membranes at defect site, connective tissue in

between

1

Cannot be evaluated because of degradation or absence of the

material

0

J Mater Sci: Mater Med (2012) 23:1951–1959 1953

123

some remnants of the PTMC membrane could still be

identified after 12 weeks (Fig. 3).

After 2 weeks both the collagen membrane and the

PTMC membrane was surrounded by a thin fibrous cap-

sule. The thin fibrous capsule around the collagen mem-

branes was composed of loose connective tissue fibers with

the presence of very few macrophages, giant cells and

other inflammatory cells. The fibrous capsule around the

PTMC membrane was also composed of loose connective

tissue fibers though appeared thicker and showed more

invasion of macrophages and giant cells compared to the

collagen treated animals. The macrophages and giant cells

appeared to erode the surface of the PTMC membrane by

phagocytosis (Fig. 2b, d).

At 2 weeks high numbers of blood vessels were present

at the tissues surrounding the PTMC membranes and

especially located in the tissues filling the defects covered

with the PTMC membranes. By contrast the animals trea-

ted with the collagen membranes showed blood vessel

formation mainly localized in and throughout the mem-

branes, as well as in the surrounding tissues.

Osteoid (bone) formation was already observed after

2 weeks in both the collagen and PTMC membrane treated

groups. Interestingly at 2 and 4 weeks, two different pat-

terns of bone formation were identified. The animals

treated with the collagen membrane tended to form osteoid

and new bone throughout the defect by forming bony islets

(Figs. 2, 3). By contrast the PTMC membrane treated

animals showed osteoid and new bone formation origi-

nating from the defect borders. The pattern of new bone

formation was found similar for both groups after

12 weeks, since the majority of the sections by then

showed complete bridging of the defect with newly formed

mature bone.

3.3 Histological analysis

Figure 4a shows the data of the histological rating of the

soft tissue response to the collagen and PTMC membrane,

respectively. Statistical analysis of these data revealed that

after 2, 4 and 12 weeks significant differences existed

between the tissue response towards the used membranes

(p \ 0.001, p \ 0.001 and p = 0.006, respectively). The

tissue reaction to the PTMC membrane shows a higher

density of inflammatory cells after 2, 4 and 12 weeks

compared to the collagen membrane. For both membranes

Fig. 1 Light micrographs of rat mandibular defects covered with either

a collagen (a, c) or a PTMC (b, d) membrane after 2 weeks of

implantation. The collagen and PTMC membrane can be readily

distinguished. Degradation of the PTMC membrane can be observed.

Toluidin blue with basic fuchsin as counterstain was used for the

staining of the histological sections. (p) polymer, (ct) connective tissue,

(b) bone, (fc) fibrous capsule, (asterisk) osteoid, (filled triangle)

collagen membrane, arrows indicate phagocytosed intracellular frag-

ments. Figures ‘c and d’ magnifications (920) of the marked regions

from respectively figures ‘a and b’, which are 92 magnifications

1954 J Mater Sci: Mater Med (2012) 23:1951–1959

123

the score increased at 12 weeks, which means that there

was a reduction in inflammatory cell density, and the

number of giant cells, and an increase in vascularization of

the surrounding tissue.

The data on tissue formation at the defect site, shown in

Fig. 4b, was similar for both membranes. After 2 and

4 weeks the majority of the sections showed amounts of

osteoid and bone formation in the defects for both mem-

brane treated groups. Between 4 and 12 weeks the newly

formed bone had matured and after 12 weeks extensive

amounts of mature bone were found for both membrane

groups. There were no statistical differences in the tissue

proliferation at the defect sites between the collagen and

PTMC treated animals (p = 0.467). Moreover, after

12 weeks mature bone with bone marrow had formed at the

defect sites, whereas after 4 weeks bone could be assessed

as immature.

Figure 4c presents the data on the space maintaining

properties of the two membranes. The PTMC membrane

showed a distinctively maintained space at the level of the

mandibular defects, between the buccal and lingual placed

membranes after 2 and 4 weeks (Figs. 1, 2). This space

was filled with either connective tissue, osteoid or newly

formed bone depending on the time of evaluation,

respectively 2, 4 or 12 weeks. The collagen membranes by

contrast showed no space maintaining at all after 2 and

4 weeks. After 12 weeks the space maintaining properties

of both membranes could not be assessed for two reasons:

firstly, because bone had bridged the majority of the

defects, and as a result there was no ‘space’ left to be

assessed, and secondly because of the extensive degrada-

tion of both membranes. The collagen membranes could

not be discerned anymore due to degradation and similarity

to the host connective tissue and for the PTMC membranes

only some phagocytosed particles were observed. As a

consequence the space maintaining properties could not be

assessed, and score ‘0’ was assigned to all specimens of the

12 weeks group.

4 Discussion

In the present study a rat model was used to investigate the

soft and hard tissue response to a barrier membrane com-

posed of solid PTMC. Furthermore the space maintaining

properties were assessed, as space maintaining is an

Fig. 2 Light micrographs of rat mandibular defects covered with

either a collagen (a, c) or a PTMC (b, d) membrane after 4 weeks of

implantation. The collagen membrane can be distinguished from the

rats connective tissue. Bone forms in and around the collagen

membrane, by contrats bone formation underneath the PTMC

membrane originates from the defect borders. Toluidin blue with

basic fuchsin as counterstain was used for the staining of the

histological sections. (p) polymer, (ct) connective tissue, (b) bone,

(fc) fibrous capsule, (asterisk) osteoid, (filled triangle) collagen

membrane, arrows indicate phagocytosed intracellular fragments.

(filled square) Indicates surface erosion degradation by macrophage

and giant cell activity. Figures ‘c and d’ magnifications (920) of the

marked regions from respectively figures ‘a and b’, which are 92

magnifications

J Mater Sci: Mater Med (2012) 23:1951–1959 1955

123

important factor in GBR. While copolymers prepared from

lactide, glycolide and TMC have been intensively investi-

gated and are now in clinical use [17–19], this is the first

time barrier membranes prepared from TMC only have

been applied in GBR.

Critical size rat mandibular defects were selected as the

orthotopic model to evaluate the PTMC membranes. Previ-

ously performed studies have shown that 5.0 mm circular rat

mandibular defects are of critical size and therefore will not

heal spontaneously during the lifetime of the animal [25, 26].

Fig. 3 Light micrographs of rat mandibular defects covered with

either a collagen (a, c) or a PTMC (b, d) membrane after 12 weeks of

implantation. Bone bridges the mandibular defects in both membrane

groups. The collagen membrane is not present anymore and has

resorbed. The PTMC membrane eroded completely, only a few

phagocytosed polymer particles are in situ. Toluidin blue with basic

fuchsin as counterstain was used for the staining of the histological

sections. (p) polymer, (ct) connective tissue, (b) bone, (fc) fibrous

capsule, (asterisk) osteoid, (v) marks blood vessel, (f) fat cell, arrowsindicate phagocytosed intracellular fragments. Figures ‘c and d’ are

magnifications (920) of the marked regions from respectively figures

‘a and b’, which are 92 magnifications

Fig. 4 a Histological scoring results for the soft tissue response to the

collagen and PTMC membrane after 2, 4 and 12 weeks. Error barsrepresent means ± standard deviation for n = 4, *p \ 0.05. b Scoring

results for the proliferated tissue at the mandibular defect site. Errorbars represent means ± standard deviation for n = 4, *p \ 0.05.

c Scoring results for the space-maintaining properties for both

membranes. At time = 12 weeks the space-maintaining properties

could not be assessed because of closure of the mandibular defects

and degradation of both the membranes. Error bars represent

means ± standard deviation for n = 4, *p \ 0.05

1956 J Mater Sci: Mater Med (2012) 23:1951–1959

123

The benign soft tissue response to PTMC membranes

indicated that PTMC membranes are promising materials

for use in GBR techniques. The thin cellular capsule

around the PTMC membranes implies that the host has

initiated the foreign body reaction that is observed with

many implant materials, and at the same time still is

involved in a response at the interface with the implanted

material. The results after 2, 4, and 12 weeks show that the

soft tissue response to the PTMC membrane is accompa-

nied by a larger influx of macrophages, giant cells and

other inflammatory cells concentrated at the tissue–implant

interface when compared to the collagen membrane.

However, this reaction can be regarded as a normal tissue

response to the PTMC, as the degradation is characterized

by a surface erosion process [22, 27]. The surface erosion

process attracted cells involved in the foreign-body reac-

tion and degradation to the tissue–implant interface. This is

in accordance with the results of previous studies, where

also an influx of macrophages and formation of foreign-

body giant cells was observed at the tissue–implant inter-

face [22, 27]. The foreign-body giant cells were only

transiently present and disappeared with increasing

implantation time. Therefore, it can be concluded that the

tissue reaction (i.e. foreign-body reaction) induced by the

PTMC membrane was mild and transient.

The tissue response to the collagen membrane was

characterized by an even milder reaction, possibly due to

its resemblance to native collagen. The degradation profile

is characterized by a macrophage and polymorphonuclear

leukocyte associated degradation as well as an enzymatic

degradation profile [28, 29].

The histological evaluation of the proliferated tissue at

the defect site showed for both membranes that increasing

amounts of bone had formed with time, witnessing the

defect closure with increasing implantation time. Recent

quantitative analysis of the amount of bone formed in rat

mandibular defects covered using PTMC and collagen (and

e-PTFE) membranes had shown that significant amounts of

bone were formed, without statistically significant differ-

ences between the membranes [16]. This implies that use

of the PTMC membrane facilitates in bone regeneration in

critical size defects in rats. These results suggest that the

PTMC membrane is biocompatible with bone and assists in

promoting bone regeneration in critical size defects.

As mentioned before, two different patterns of bone

formation were observed. The bone formed beneath the

PTMC membranes tended to grow from the defect borders

towards the centre of the defect. This way of defect closure

has been described often in literature and seems to be the

most common in the formation of new bone when non-

osteoinductive biomaterials are applied [30–32]. The col-

lagen membrane showed bone formation within the mem-

brane and throughout the defect originating from bony

islets and the defect border. This has been previously

described by Hoogeveen et al. in a comparative study

between three different barrier membranes used in GBR

[33]. A possible explanation for the formation of these

bony islets could be that the animal derived natural colla-

gen has osteoinductive properties. Nonetheless, the differ-

ent bone formation patterns did not show statistical

differences regarding the tissue proliferation at the defect

site between the two membranes for the different time

periods. After just 2 weeks osteoid formation was present

for both membranes, again indicating that the PTMC is a

bone biocompatible material that allows for bone formation

in the space in between the membranes.

Space maintaining properties of the barrier membrane

are important in GBR to lead to predictable bone forma-

tion, by providing space for the blood clot and new bone

formation in the defect. During the implantation procedure

it became clear that the handling and space maintaining

properties of 0.3 mm thick PTMC membranes were supe-

rior to the collagen membrane. This was confirmed by the

results of the histological evaluation after 2 and 4 weeks.

Compared to the collagen membrane the PTMC membrane

exerted better space maintaining properties.

The space maintaining properties of the membranes

after 12 weeks could not be assessed for two reasons.

Firstly, since both the PTMC and collagen membrane had

almost completely resorbed, and therefore had lost their

mechanical strength resulting in a complete loss of space

maintaining properties. A second reason is that new bone

had bridged the majority of the defects and space main-

taining properties cannot be assessed without a distinct

defect in the rat mandible.

An important problem with collagen membranes is that

almost immediately upon implantation they loose their

mechanical strength and, therefore, their space maintaining

properties. The collagen membrane is therefore best used in

combination with (autologous) grafting materials to

achieve comparable results to non-resorbable e-PTFE

barrier membranes [20, 34–36]. The better mechanical

properties of the resorbable PTMC membrane should

enable comparable results to those of non-resorbable

e-PTFE membrane without the use of grafting materials.

Before conducting clinical trials in humans, this should

preferably be evaluated in a next study where deeper

defects with larger volumes are created. This will empha-

size the importance of the space maintaining properties of

the membranes. A model such as the one described by Oh

et al. [32] would be useful as they created buccal dehis-

cence defects in the jaw bone around dental implants

measuring 4 mm in height from the crestal bone, 3 mm in

depth from the surface and 3 mm in width in dogs. In this

model the membranes space maintaining properties can be

better assessed as larger volumes of deficient bone are

J Mater Sci: Mater Med (2012) 23:1951–1959 1957

123

involved, also its biological behaviour around dental

implants and more importantly its behaviour towards

exposure to the oral environment can be assessed. Never-

theless, the present study demonstrated histologically that

the PTMC membrane allows bone growth and therefore

makes it suitable for use in GBR techniques.

Finally, the general opinion in medicine is that

(resorbable) animal derived materials should be replaced by

similarly performing (resorbable) synthetic materials when

available, in order to avoid possible transmission of disease

[3–5]. In this respect, the application of PTMC as a barrier

membrane in GBR techniques could be a step forward as an

alternative for the animal derived collagen membranes.

Acknowledgments Gratitude is expressed to Ms. M. B. M. van

Leeuwen for her assistance and advice during the histological pro-

cedures. Ms. Y. Heddema and Ms. N. Broersma are acknowledged for

their assistance during the surgical procedures. Furthermore we would

like to thank Geistlich Biomaterials for provision of the Geistlich

BioGide regenerative membranes.

Open Access This article is distributed under the terms of the

Creative Commons Attribution License which permits any use, dis-

tribution, and reproduction in any medium, provided the original

author(s) and the source are credited.

References

1. Hollinger JO, Buck DC, Bruder SP. Biology of bone healing: its

impact on clinical therapy. In: Lynch SE, Genco RJ, Marx RE,

editors. Tissue engineering. Applications in maxillofacial surgery

and periodontics. Chicago: Quintessence; 1999. p. 17–53.

2. Kay SA, Wisner-Lynch L, Marxer M, Lynch SE. Guided bone

regeneration: integration of a resorbable membrane and a bone

graft material. Practical periodontics and aesthetic dentistry.

PPAD. 1997;9:185–96.

3. von Arx T, Cochran DL, Schenk RK, Buser D. Evaluation of a

prototype trilayer membrane (PTLM) for lateral ridge augmen-

tation: an experimental study in the canine mandible. Int J Oral

Maxillofac Surg. 2002;31:190–9.

4. Pauli G. Tissue safety in view of CJD and variant CJD. Cell

Tissue Bank. 2005;6:191–200.

5. Fishman JA. Infection in xenotransplantation. J Card Surg.

2001;16:363–73.

6. Bergsma JE, Debruijn WC, Rozema FR, Bos RRM, Boering G.

Late degradation tissue-response to poly(L-lactide) bone plates

and screws. Biomaterials. 1995;16:25–31.

7. Bostman OM. Osteolytic changes accompanying degradation of

absorbable fracture fixation implants. J Bone Jt Surg. 1991;73:

679–82.

8. Bostman OM. Absorbable implants for the fixation of fractures.

J Bone Jt Surg. 1991;73A:148–53.

9. Bostman O, Partio E, Hirvensalo E, Rokkanen P. Foreign-body

reactions to polyglycolide screws—observations in 24/216 mal-

leolar fracture cases. Acta Orthop Scand. 1992;63:173–6.

10. Bostman OM, Pihlajamaki HK, Partio EK, Rokkanen PU. Clin-

ical biocompatibility and degradation of polylevolactide screws

in the ankle. Clin Orthop. 1995;320:101–9.

11. Hurzeler MB, Quinones CR, Hutmacher D, Schupbach P. Guided

bone regeneration around dental implants in the atrophic alveolar

ridge using a bioresorbable barrier—an experimental study in the

monkey. Clin Oral Implants Res. 1997;8:323–31.

12. Tams J, Rozema FR, Bos RRM, Roodenburg JLN, Nikkels PGJ,

Vermey A. Poly(L-lactide) bone plates and screws for internal

fixation of mandibular swing osteotomies. Int J Oral Maxillofac

Surg. 1996;25:20–4.

13. von Arx T, Kurt B, Hardt N. Treatment of severe peri-implant

bone loss using autogenous bone and a resorbable membrane—

case report and literature review. Clin Oral Implants Res.

1997;8:517–26.

14. Wu LB, Ding JD. In vitro degradation of three-dimensional

porous poly(D,L-lactide-co-glycolide) scaffolds for tissue engi-

neering. Biomaterials. 2004;25:5821–30.

15. Taylor MS, Daniels AU, Andriano KP, Heller J. Six bioabsorb-

able polymers—in vitro acute toxicity of accumulated degrada-

tion products. J Appl Biomater. 1994;5:151–7.

16. Van Leeuwen AC, Huddleston Slater JJR, Gielkens PFM, De

Jong JR, Grijpma DW, Bos RRM. Guided bone regeneration in

rat mandibular defects using resorbable poly(trimethylene car-

bonate) barrier membranes. Acta Biomater. 2012;8:1422–9.

17. Stavropoulos F, Dahlin C, Ruskin J, Johansson C. A comparative

study of barrier membranes as graft protectors in the treatment of

localized bone defects—an experimental study in a canine model.

Clin Oral Implants Res. 2004;15:435–42.

18. Amecke B, Bendix D, Entenmann G. Synthetic resorbable

polymers based on glycolide, lactides, and similar monomers. In:

Wise DL, editor. Encyclopedic handbook of biomaterials and

bioengineering. New York: Marcel Dekker Inc.; 1995. p. 982–3.

19. Zwahlen RA, Cheung LK, Zheng L, Chow RLK, Li T, Schukn-

echt B, Graetz KW, Weber FE. Comparison of two resorbable

membrane systems in bone regeneration after removal of wisdom

teeth: a randomized-controlled clinical pilot study. Clin Oral

Implants Res. 2009;20:1084–91.

20. Donos N, Kostopoulos L, Karring T. Alveolar ridge augmentation

using a resorbable copolymer membrane and autogenous bone

grafts—an experimental study in the rat. Clin Oral Implants Res.

2002;13:203–13.

21. Pego AP, Grijpma DW, Feijen J. Enhanced mechanical properties

of 1,3-trimethylene carbonate polymers and networks. Polymer.

2003;44:6495–504.

22. Pego AP, Van Luyn MJA, Brouwer LA, van Wachem PB, Poot

AA, Grijpma DW, Feijen J. In vivo behavior of poly(1,3-tri-

methylene carbonate) and copolymers of 1,3-trimethylene car-

bonate with D,L-lactide or epsilon-caprolactone: degradation and

tissue response. J Biomed Mater Res Part A. 2003;67A:1044–54.

23. Zhang Z, Kuijer R, Bulstra SK, Grijpma DW, Feijen J. The in

vivo and in vitro degradation behavior of poly(trimethylene

carbonate). Biomaterials. 2006;27:1741–8.

24. Gielkens PFM, Schortinghuis J, de Jong JR, Raghoebar GM,

Stegenga B, Bos RRM. Vivosorb (R), Bio-Gide (R), and Gore-

Tex (R) as barrier membranes in rat mandibular defects: an

evaluation by microradiography and micro-CT. Clin Oral

Implants Res. 2008;19:516–21.

25. Schortinghuis J, Ruben JL, Meijer HJA, Bronckers ALJJ,

Raghoebar GM, Stegenga B. Microradiography to evaluate bone

growth into a rat mandibular defect. Arch Oral Biol. 2003;48:

155–60.

26. Kaban LB, Glowacki J, Murray JE. Repair of experimental

mandibular bony defects in rats. Surg Forum. 1979;30:519–21.

27. Bat E, Plantinga JA, Harmsen MC, van Luyn MJA, Feijen J,

Grijpma DW. In vivo behavior of trimethylene carbonate and

epsilon-caprolactone-based (co)polymer networks: degradation

and tissue response. J Biomed Mater Res Part A. 2010;95A:

940–9.

28. von Arx T, Broggini N, Jensen SS, Bornstein MM, Schenk RK,

Buser D. Membrane durability and tissue response of different

1958 J Mater Sci: Mater Med (2012) 23:1951–1959

123

bioresorbable barrier membranes: a histologic study in the rabbit

calvarium. Int J Oral Maxillofac Implants. 2005;20:843–53.

29. Kozlovsky A, Aboodi G, Moses O, Tal H, Artzi Z, Weinreb M,

Nemcovsky CE. Bio-degradation of a resorbable collagen mem-

brane (Bio-Gide((R))) applied in a double-layer technique in rats.

Clin Oral Implants Res. 2009;20:1116–23.

30. Mueller AA, Rahn BA, Gogolewski S, Leiggener CS. Early dural

reaction to polylactide in cranial defects in rabbits. Pediatr

Neurosurg. 2005;41:285–91.

31. Mokbel N. Bou Serhal C, Matni G, Naaman N. Healing patterns

of critical size bony defects in rat following bone graft. Oral

Maxillofac Surg. 2008;12:73–8.

32. Oh TJ, Meraw SJ, William EJ, Giannobile WV, Wang HL. Com-

parative analysis of collagen membranes for the treatment of implant

dehiscence defects. Clin Oral Implants Res. 2003;14:80–90.

33. Hoogeveen EJ, Gielkens PFM, Schortinghuis J, Ruben JL,

Huysmans M-DNJM, Stegenga B. Vivosorb (R) as a barrier

membrane in rat mandibular defects. An evaluation with trans-

versal microradiography. Int J Oral Maxillofac Surg. 2009;38:

870–5.

34. Lundgren AK, Lundgren D, Sennerby L, Taylor A, Gottlow J,

Nyman S. Augmentation of skull bone using a bioresorbable

barrier supported by autologous bone grafts—an intra-individual

study in the rabbit. Clin Oral Implants Res. 1997;8:90–5.

35. Lundgren AK, Sennerby L, Lundgren D, Taylor A, Gottlow J,

Nyman S. Bone augmentation at titanium implants using autol-

ogous bone grafts and a bioresorbable barrier—an experimental

study in the rabbit tibia. Clin Oral Implants Res. 1997;8:82–9.

36. Simion M, Misitano U, Gionso L, Salvato A. Treatment of

dehiscences and fenestrations around dental implants using

resorbable and nonresorbable membranes associated with bone

autografts: a comparative clinical study. Int J Oral Maxillofac

Implants. 1997;12:159–67.

J Mater Sci: Mater Med (2012) 23:1951–1959 1959

123